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Addressing Shewanella oneidensis "cytochromome": the first step towards high-throughput expression of cytochromes c.

Abstract：

:Integrated studies that address proteins structure and function in the new era of systems biology and genomics often require the application of high-throughput approaches for parallel production of many different purified proteins from the same organism. Cytochromes c-electron transfer proteins carrying one or more hemes covalently bound to the polypeptide chain-are essential in most organisms. However, they are one of the most recalcitrant classes of proteins with respect to heterologous expression because post-translational incorporation of hemes is required for proper folding and stability. We have addressed this challenge by designing two families of vectors (total of 6 vectors) suitable for ligation-independent cloning and developing a pipeline for expression and solubility analysis of cytochromes c. This system has been validated by expression analysis of thirty genes from Shewanella oneidensis coding for cytochromes c or cytochromes c-type domains predicted to have 1-4 hemes. Out of 30 targets, 26 (87%) were obtained in soluble form in one or more vectors. This work establishes a methodology for high-throughput expression of this class of proteins and provides a clone resource for the microbiological and functional genomics research communities.

journal_name

Protein Expr Purif

journal_title

Protein expression and purification

authors

Londer YY,Giuliani SE,Peppler T,Collart FR

doi

10.1016/j.pep.2008.06.014

subject

Has Abstract

pub_date

2008-11-01 00:00:00

pages

128-37

issue

1

eissn

1046-5928

issn

1096-0279

pii

S1046-5928(08)00169-1

journal_volume

62

pub_type

杂志文章

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